PROJECT SUMMARY The recent increase in obesity is a major public health concern. Since energy balance regulation is coordinated by communication between the gastrointestinal (GI) tract and the brain, understanding these gut-brain interactions will enable the development of novel obesity treatments. The hypothalamus and the hindbrain are critical brain regions that integrate information from the gut to control food intake. Here, I will leverage recent technological advances to explore the regulation of both these brain regions in awake, behaving animals. Within the hypothalamus, agouti-related protein (AgRP)-expressing neurons are essential for food intake control. Activity in AgRP neurons is high during hunger and is rapidly inhibited by food. My recent work demonstrates that AgRP neurons are primarily regulated by calorie intake, rather than sensory detection of food, since direct gastric infusion of macronutrients into the stomach rapidly suppresses AgRP neuron activity in vivo. Further, this effect is recapitulated by administration of GI satiation signals normally released following food consumption. However, the mechanisms through which the gut transmits signals to AgRP neurons remain unknown. The mentored phase (Aims I and II) of this grant will build upon my previous work by elucidating the mechanisms through which nutrient detection in the gut leads to AgRP neuron activity reductions. Specifically, these aims will determine whether GI signals are transmitted vagally or through direct action on the brain, and will uncover the AgRP axon projections that transmit nutritive signals throughout the brain. Importantly, the mentored experiments will afford me training in peripheral manipulation of the GI tract, as well as in vivo calcium imaging of individual neurons using microendoscopy and 2-photon microscopy, expanding my technical expertise and enabling the proposed R00 experiments. The hindbrain nucleus tractus solitarius (NTS) is the first central site of integration of GI-derived signals from vagal afferents, and is a key signaling node that transmits signals from the gut to higher-order brain structures such as the hypothalamus. For the independent phase (Aims III and IV) of my grant, I have designed experiments that build upon both my graduate and postdoctoral training to determine how different hindbrain NTS neuron populations receive signals from the GI tract, at unprecedented levels of temporal and cellular detail. These complementary research aims combined with the proposed career development activities will provide me with the training necessary to successfully transition to independence, under the guidance of my mentorship team who have extensive collective experience with neuroscience techniques and mentorship. Overall, this award will facilitate my career as an independent investigator characterizing the role of gut-brain signaling on the in vivo activity dynamics of feeding-relevant neurons.